Abstract Arthropod-borne viruses represent a major threat to humans on a global scale, and of these flaviviruses transmitted by Aedes aegypti mosquitoes are the most significant risk to human health. Understanding the immunologic events that take place in the human host, particularly in the skin where virus is transmitted during mosquito feeding, represents a major gap in our understanding of arbovirus disease. Addressing this gap is a central theme of the Funding Opportunity PAR-18-860, ?Immune Response to Arthropod Blood Feeding?, and is the goal of the current proposal. To best define these events, we propose a transdisciplinary study between the Barratt-Boyes lab, with expertise in human skin immunology, and the Vasilakis lab, with expertise in arbovirology and mosquito-virus-host interactions. The Barratt-Boyes group has established an ex vivo model of dengue (DENV) and Zika virus (ZIKV) infection in human skin using large-area explants from anonymous healthy donors, and have defined the immunologic events that occur in skin in the absence of mosquito probing. The unique access of the Vasilakis group to insectary facilities approved for working with live Aedes spp. infected with DENV or ZIKV allows us to now explore the contribution of the mosquito to viral pathogenesis. Thus, we will bring together three essential components ? live mosquitoes, human skin, and pathogenic viruses ? to address for the first time the immunologic events that occur during vector-borne transmission of flaviviruses in human skin. This is a truly innovative proposal; to our knowledge, no publications exist describing the response of human skin to pathogenic flaviviruses transmitted by live mosquitoes, and no other research group has an experimental system such as ours to fill this research gap. We have two specific aims: (1) what is the immune response of human skin to mosquito probing in the absence of virus, and (2) how does mosquito probing affect DENV and ZIKV replication and spread in human skin. The results of our study will have a strong and lasting impact on the field of immunology and transmission of vector-borne pathogens. The work will establish the ex vivo mosquito-skin-virus system as a foundation for future studies on pathogenesis, therapy and vaccine development.